During carcinogenesis, alterations occur in the biosynthesis of carbohydrate structures on the cell surface, and several different carbohydrates linked either to proteins or to lipids have been recognized to be tumor-associated antigens. TF-Antigen (TF-Ag) is a tumor-associated antigen of several carcinomas, including breast, lung, bladder, prostate and pancreas (1-4). TF-Ag has been proposed to be involved in the metastatic process (5-10).
TF-Ag is Galactoseβ1-3 N-Acetyl Galactosamine, a disaccharide attached to a protein by an alpha O-serine or O-threonine linkage. TF-Ag was discovered by Thomsen, Friedenreich and Hueber in the late 1920's (2). TF-Ag is hidden on various normal cell membranes because it is linked to other carbohydrates, either by tertiary structures or by highly negative-charged sialic acid (3,7). The densities of TF-Ag predict the histologic grade of carcinomas (9,11), the invasive potential, and the probability of early recurrence in breast (2,12-13), urinary bladder, and prostatic carcinoma (14).
TF-Ag is more than an immunopathologic marker; it is postulated to have a role in adhesion and metastasis (2). Reversible or irreversible adhesion is a primary step of invasion (15) and may occur when the TF-Ag adhesion molecules recognize ligands such as galectins or other lectins (4,16). There is an increased expression of TF-Ag in metastatic tumors, and lectins that bind TF-Ag are in common sites of metastatic tumor growth (17). Ligands for TF-Ag adhesion have been found in the vascular endothelium, the liver, the bone marrow and the lymph nodes (18-19) and this may explain how TF-Ag levels are related to carcinoma aggressiveness (20).
There have been reports concerning the immunotherapeutic value of an induced immune response to carbohydrate tumor associated antigens (8,26-29). For example, O'Boyle et al show development of very low level IgG and IgM responses to two antigens which are related to TF-Ag, Tn (GalNAc) and sialylated Tn (NANAα2-6GalNAc) in colon cancer patients (26-27). Longenecker's studies in breast cancer patients used a keyhole limpet hemocyanin (KLH) conjugate of sialyl Tn and obtained higher responses and observed some clinical response in these patients (29-30). Springer and Desai used vaccination with a T/Tn vaccine composed of types O and MN red blood cell derived glycoproteins which resulted in improved breast cancer patient survival although only small amounts of IgM were made (15). Immunization of breast cancer patients with Globo H (a hexasaccharide which contains TF-Ag), conjugated to KLH injected with the adjuvant QS21 to improve immunogenicity resulted in most patients forming only an IgM response (31). However, IgM is generally of lower affinity and specificity than IgG, and represents a less mature immune response, and many previous studies relating to antibodies to TF-Ag involve IgM antibodies. Further, some anti-TF-Ag antibodies are not clinically useful because they cause undesirable proliferation of tumor cells (45). Moreover, while peptides which can bind to TF-Ag and inhibit cell adhesion in vitro have been described (5), it has also been found that some peptides which can bind to the TF-Ag on human carcinoma cell lines and interfere with cell aggregation exhibit a low monomeric affinity for the TF-Ag, and therefore cannot be dissolved at high concentration in aqueous buffers, which significantly limit their potential use (Landon et al., Journal of Protein Chemistry, (2003) Vol. 22: pp 193-204).
One study purported to investigate the effect of a monoclonal antibody to TF-Ag on cancer cells in a mouse model (Shigeoka, et al. Tumor Biology (1999) 20: 138-146). However, that study attempted to simulate metastasis by direct injection into the liver of cancer cells that were pre-incubated with the monoclonal antibody before being injected. Nodule formation in the livers of mice injected with the cells pre-incubated with the monoclonal antibody were compared to mice receiving injections with cancer cells that were not pre-incubated with the monoclonal antibody. While there were fewer nodules in mice receiving the pre-incubated cells, this study did not provide a relevant model of antibody-mediated inhibition of metastasis because, due to the pre-incubation of the cancer cells with the monoclonal antibody, there was no requirement for the antibody to travel through the body to locate and bind to cells expressing the TF-Ag. Further, there was no assessment as to whether the nodule formation was the result of cancer cells traveling through the endothelium in a manner similar to the natural metastatic process or whether the nodules were simply the result of lodging and proliferation of the injected cells.
Finally, while the production and characterization of another anti TF-Ag monoclonal antibody has been described (21), none of the aforementioned studies demonstrate that the use of a monoclonal antibody to TF-Ag would inhibit either metastasis or growth of cancer cells in an individual.